The present invention is in the field of enzyme proteins that are related to the synthase enzyme subfamily, recombinant DNA molecules, and protein production. The present invention specifically provides a novel alternative splice form of a squalene synthase enzyme and nucleic acid molecules encoding the novel splice form, all of which are useful in the development of human therapeutics and diagnostic compositions and methods.
Many human enzymes serve as targets for the action of pharmaceutically active compounds. Several classes of human enzymes that serve as such targets include helicase, steroid esterase and sulfatase, convertase, synthase, dehydrogenase, monoxygenase, transferase, kinase, glutanase, decarboxylase, isomerase and reductase. It is therefore important in developing new pharmaceutical compounds to identify target enzyme proteins that can be put into high-throughput screening formats. The present invention advances the state of the art by providing novel human drug target enzymes related to the synthase subfamily.
Synthases
The novel human protein, and encoding gene, provided by the present invention is related to the family of synthases in general and squalene synthases (also known by such names as squalene synthetases and farnesyl-diphosphate farnesyltransferases) in particular. Furthermore, the protein of the present invention may be a novel isoform of the protein/gene provided in Genbank gi4758350. Specifically, the protein/cDNA of the present invention differs from the art-known protein of gi4758350 in that the fourth exon is spliced out of the protein/cDNA of the present invention (see the amino acid sequence alignments in FIG. 2).
Squalene synthase is important for catalyzing the first specific step in the biosynthesis of cholesterol, which is the conversion of trans-farnesyldiphosphate to squalene. Squalene synthase regulates this major control point in sterol, as well as isoprene, biosynthesis in eukaryotes (Robinson et al., Mol Cell Biol 1993 May;13(5):2706-17). Squalene synthase thus occupies a critical regulatory position in cholesterol synthesis (Schechter et al., Genomics 1994 Mar. 1;20(1):116-8). Furthermore, the squalene synthase gene has been associated with Rec syndrome (Patterson et al., Am. J. Hum. Genet. 57: A91 only, 1995).
Loss of promoter activity and response to sterols has been localized to a 69-bp region that is positioned 131 bp upstream from the transcription start site; this region contains a sterol regulatory element-1, which is found in other sterol regulated genes, and two possible NF1 binding sites (Guan et al., Biol. Chem. 270: 21958-21965, 1995).
For a further review of squalene synthases, see McKenzie et al., J Biol Chem 1992 Oct. 25;267(30):21368-74; Summers et al., Gene 1993 Dec. 22;136(1-2Che):185-92; Jiang et al., J Biol Chem 1993 Jun. 15;268(17):12818-24; and Soltis et al., Arch Biochem Biophys 1995 Feb. 1;316(2):713-23.
Due to their importance in cholesterol biosynthesis, novel human squalene synthase proteins/genes, such as provided by the present invention, are valuable as potential targets for the development of therapeutics to treat cholesterol-related diseases/disorders such as cardiovascular diseases. Furthermore, SNPs in squalene synthase genes, such as provided by the present invention, may serve as valuable markers for the diagnosis, prognosis, prevention, and/or treatment of cholesterol-related diseases/disorders such as cardiovascular diseases.
Using the information provided by the present invention, reagents such as probes/primers for detecting the SNPs or the expression of the protein/gene provided herein may be readily developed and, if desired, incorporated into kit formats such as nucleic acid arrays, primer extension reactions coupled with mass spec detection (for SNP detection), or TaqMan PCR assays (Applied Biosystems, Foster City, Calif.).
Enzyme proteins, particularly members of the synthase enzyme subfamily, are a major target for drug action and development. Accordingly, it is valuable to the field of pharmaceutical development to identify and characterize previously unknown members of this subfamily of enzyme proteins. The present invention advances the state of the art by providing previously unidentified human enzyme proteins, and the polynucleotides encoding them, that have homology to members of the synthase enzyme subfamily. These novel compositions are useful in the diagnosis, prevention and treatment of biological processes associated with human diseases.
The present invention is based in part on the identification of amino acid sequences of human enzyme peptides and proteins that are related to the synthase enzyme subfamily, as well as allelic variants and other mammalian orthologs thereof. These unique peptide sequences, and nucleic acid sequences that encode these peptides, can be used as models for the development of human therapeutic targets, aid in the identification of therapeutic proteins, and serve as targets for the development of human therapeutic agents that modulate enzyme activity in cells and tissues that express the enzyme. Experimental data as provided in FIG. 1 indicates expression in humans in placenta, T-cells from T-cell leukemia, fetal brain, pancreas, Burkitt lymphoma, bladder, and liver.